Abstract
Abstract Metastasis Suppressor Genes (MSGs) are able to inhibit metastasis without reducing primary tumor size. Nm23 was the first MSGs to be identified. The biochemical basis for the anti-metastatic effect of Nm23 has been subject of many studies but remains unresolved. We performed co-immunoprecipitation assays using flag-tagged constructs of both the human and murine forms of Nm23 (Nm23-H1 and Nm23-M1, respectively) over-expressed in the mouse mammary carcinoma cell line 4T1. Co-immunoprecipitation assays were also performed on tissue extracts of primary tumors and metastases obtained from Balb/C mice injected with these cell lines. Using Mass-Spectrometry, new binding partners were identified including the actin-binding protein Gelsolin. Gelsolin is an actin regulatory protein that modulates actin assembly and disassembly, and is believed to regulate cell motility in vivo through modulation of the actin network. Recently, Gelsolin has been added to the list of MSGs. An interaction between Gelsolin and Nm23 was validated in a murine cell line (4T1- Nm23-H1/M1 clones) and in a human breast carcinoma cell lines (MCF-7). In the low- metastatic cells MCF-7 an interaction between endogenous Nm23-H1 and Gelsolin was observed. However, in the highly metastatic MDA-MB-435 and MDA-MB-231 cell lines the interaction was observed only after the over-expression of Nm23-H1. These results suggest that an Nm23-Gelsolin interaction may only take place when relatively high levels of Nm23 are expressed. We conclude that the interaction between Nm23-H1 and Gelsolin occurs in the non-metastatic state, where high levels of both proteins are present, and this interaction can contribute to the anti-metastatic property associated to Nm23. Using cultures of MDA-MB-231 stably overexpressing Gelsolin, Nm23-H1 or both proteins, the biological role of this interaction was analyzed. The two metastasis suppressors, Nm23-H1 and Gelsolin, when overexpressed in the single stable clone did not affect the cell growth; however they cooperated in decreasing the cell growth. This is the first time overexpression of Nm23-H1 has decreased cell proliferatioin and suggests that combinations of MSGs may have distinct phenotypic effects. Moreover, overexpression of both Nm23-H1 and Gelsolin in MDA-MB-231 cells reduced cell migration and cell invasion compared to the clone overexpressing the empty vector or single suppressor genes. In summary, Nm23-H1 has been demonstrated to bind another Metastasis Suppressor, Gelsolin. Additive and synergistic effects of multiple Metastasis Suppressors were identified. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4753. doi:10.1158/1538-7445.AM2011-4753
Published Version
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